CN106967765A - The fermentation process of high concentration Cordyceps sinensis polysaccharide - Google Patents
The fermentation process of high concentration Cordyceps sinensis polysaccharide Download PDFInfo
- Publication number
- CN106967765A CN106967765A CN201710355633.8A CN201710355633A CN106967765A CN 106967765 A CN106967765 A CN 106967765A CN 201710355633 A CN201710355633 A CN 201710355633A CN 106967765 A CN106967765 A CN 106967765A
- Authority
- CN
- China
- Prior art keywords
- cordyceps
- fermentation
- culture
- cordyceps militaris
- polysaccharide
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/04—Polysaccharides, i.e. compounds containing more than five saccharide radicals attached to each other by glycosidic bonds
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/14—Fungi; Culture media therefor
Landscapes
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Health & Medical Sciences (AREA)
- Wood Science & Technology (AREA)
- Genetics & Genomics (AREA)
- Biotechnology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Health & Medical Sciences (AREA)
- General Engineering & Computer Science (AREA)
- Biochemistry (AREA)
- Microbiology (AREA)
- Medicinal Chemistry (AREA)
- Mycology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Biomedical Technology (AREA)
- Virology (AREA)
- Botany (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Medicines Containing Plant Substances (AREA)
Abstract
The present invention provides a kind of fermentation process of high concentration Cordyceps sinensis polysaccharide, greatly optimize the process conditions of Polysaccharides in Cultured Cordyceps militaris fermentation, effectively shorten fermentation period and production cost, the concentration for the Cordyceps sinensis polysaccharide that the overall production efficiency of raising and fermenting and producing go out, is that the offer of the fermenting and producing of modern industrialization Cordyceps sinensis polysaccharide is provided powerful support for.
Description
Technical field
The invention belongs to bio-fermentation engineering field, and in particular to a kind of fermentation process of high concentration Cordyceps sinensis polysaccharide.
Background technology
Cordyceps militaris is Ascomycotina Ascomycota, Hypocreales Hypocreales, Clavicipitaceae
Clavicipitaceae, Cordyceps (Cordyceps) type sepecies.Scientific name is Cordyceps militaris, also known as
Northern Chinese caterpillar Fungus, northern Cordyceps militaris, cordyceps sinensis etc., by stroma (i.e. careless part) and sclerotium (i.e. the corpse part of worm) two parts group
Into complex, worldwide distribution but natural resources quantity is seldom.The traditional Chinese medical science thinks that cordyceps sinensis enters lung kidney two warp, can tonifying lung it is cloudy, again
Energy kidney-replenishing, cures mainly kidney deficiency, soreness of waist and knee joint, eak after being ill, chronic cough phlegm blood, night sweat etc., is that unique one kind can be while balancing, adjusting
Save the Chinese medicine of negative and positive.Substantial amounts of modern pharmacological research shows that Cordyceps militaris not only has special nutritive value, and has obvious
Medical value.Wherein Cordyceps sinensis polysaccharide be that cordyceps sinensis in-vivo content is most abundant, one of most important bioactive substance, because with anti-swollen
Knurl, anti-oxidant, radioresistance and raising body's immunity, the adrenal effect of promotion, the secretion of increase human insulin, reduction
The extensive pharmacological action such as blood sugar level of diabetes patient and receive much attention.Cordyceps sinensis polysaccharide is a kind of highly branched galactomannan
Glycan, being capable of activated macrophage stimulation antibody generation, promotion lymphocyte transformation, the antibody content and machine of raising serum IgG
The immunologic function of body, enhancing body itself anticancer presses down the ability of cancer, and clinic has been used for treating malignant tumour.In addition, Cordyceps sinensis polysaccharide
It can also improve respiratory system, anti-arrhythmia, resist myocardial ischemia, expand peripheral vascular, decompression, reducing blood lipid, suppress platelet aggregation
Collection etc..It is rapid to the demand growth of Cordyceps militaris and Cordyceps sinensis polysaccharide in current world wide, and Cordyceps sinensis polysaccharide in wild Cordyceps militaris
Content it is not high, along with the madness of people is plucked, price is increasingly expensive, and the extensive use of Polysaccharides in Cultured Cordyceps militaris is by medicine source resource
Serious restriction, and approach complex process by chemical synthesis and yield is extremely low.Pupa is produced using the method for liquid fermentation
Cordyceps sinensis polysaccharide has the advantages that with short production cycle, labor saving, small by external environment influence, and research is found from Cordyceps militaris
The Cordyceps sinensis polysaccharide obtained in fructification, mycelium and zymotic fluid is substantially identical on biochemical structure, therefore is considered as one
Plant the effective production method for substituting and polysaccharide being extracted from Natural C.militaris.But the fermentation level of current Polysaccharides in Cultured Cordyceps militaris is not high, production
The relatively low development for limiting its industrialized production of amount.It is many to Cordyceps militaris both at home and abroad compared with the numerous studies in fungi polysaccharide field
The research report of sugar is very few, and focuses mostly in recent years, is concentrated mainly on fermentation and product extracts the techniques such as separation
In aspect.
Such as patent document CN101067005 (application number 200710052357), one kind rice production cordyceps sinensis is disclosed many
Sugared method, it includes dispensing, inoculation, culture, obtains packaging step after Cordyceps sinensis polysaccharide and irradiation sterilization.In the dispensing, each original
The proportioning of material is：By weight, rice 45.0-49.0%, corn and soya-bean cake mixed powder 0.9-2%, nutrient solution 49.0-
54.1%；Nutrient solution prescription be the-220g of the potato 180 ,-22g of the white sugar 18 ,-18g of peptone 13, potassium dihydrogen phosphate 1.5-
The 2.2g, -1.2g of the magnesium sulfate 0.8, -1100g of water 900.At present, not yet there is the angle analysis from Polysaccharides in Cultured Cordyceps militaris biological metabolism
Research, the raising to Polysaccharides in Cultured Cordyceps militaris fermentation level is limited, and fermentation period is long, and cost is high, and overall production efficiency is relatively low, hair
The Cordyceps sinensis polysaccharide concentration that ferment is produced is not high enough, the need for can't far meeting modern industrialization fermenting and producing.
The content of the invention
In view of this, the present invention provides a kind of fermentation process of high concentration Cordyceps sinensis polysaccharide, greatly optimizes Cordyceps militaris many
The process conditions of sugar fermentation, effectively shorten fermentation period and production cost, improve overall production efficiency and fermenting and producing
The concentration of the Cordyceps sinensis polysaccharide gone out, is that the offer of the fermenting and producing of modern industrialization Cordyceps sinensis polysaccharide is provided powerful support for.
The technical scheme is that：A kind of fermentation process of high concentration Cordyceps sinensis polysaccharide, comprises the following steps：
（1）Cordyceps militaris spawn is inoculated into PD liquid fermentation mediums, activation culture is carried out, then by percentage by volume 10-
15% inoculum concentration is connected in seed fermentation culture medium, carries out seed culture, and seed culture condition is：20-25 DEG C of temperature, shaking table
Culture 2-4 days, obtains seed liquor；
（2）By step（1）Seed liquor is made to be inoculated in liquid fermentation medium by 5-10 % volume ratios, in 20-25 DEG C of temperature
Under conditions of, then liquid fermentation and culture 1-6 days adds active-fermented broth, makes the concentration for splitting kettle algae fermentation protein product be
0.15-0.85 mg/mL, then proceed to culture 2-9 days, and separation obtains cordyceps mycelium and zymotic fluid；
（3）From step（2）Cordyceps militaris exocellular polysaccharide is extracted in obtained zymotic fluid, from step（2）Obtained cordyceps mycelium
Middle extraction Cordyceps militaris intracellular polyse, mixing Cordyceps militaris exocellular polysaccharide and Cordyceps militaris intracellular polyse, produce Cordyceps sinensis polysaccharide；
The step（1）In seed fermentation culture medium, every liter of component is as follows：
3 g glucose, 1.5 g yeast powder supernatants, 0.1 g MgSO4,0.03 g CaCl2,0.1 g KH2PO4,0.1
GK2HPO4,2-5 diameter 3-6 mm bead；
The step（2）In liquid fermentation medium, every liter of component is as follows：
3 g lactose, 2 g molasses, 2 g bean cake powders, 1.5 g peptones, 0.1 g MgSO4,0.03 g CaCl2,0.1 g
KH2PO4、0.1 g K2HPO4 ；
The active-fermented broth is splits kettle algae tunning, and its preparation method is：
Over cleaning of learning from else's experience splits kettle algae raw material, sets enzymatic hydrolysis condition as solid-liquid ratio 1: 10（ m/V）, cellulase Cellulase
ACCF-4740 additions 2%, 55 DEG C of temperature, pH 4.5, the h of reaction time 1.3, are digested, cellulose hydrolyzation terminates
Afterwards, boiling water bath goes out enzyme 15 min, and 9000 r/min centrifugations collect supernatant and obtain splitting kettle algae cellulose hydrolyzation liquid；Split kettle algae
Enzymolysis liquid goes out after enzyme activity, regulation pH value to 6.2-6.6, high pressure steam sterilization（121 DEG C, 20 min）, then it is inoculated with volume fraction
The Pediococcus pentosaceus liquid of 1% MRS culture mediums activation stands, fermented to kettle algae enzymolysis liquid, 37 DEG C of constant incubators are split；
After fermentation ends, 9000 r/min centrifugations collect supernatant and obtain splitting kettle algae tunning, the active-fermented broth as prepared.
Further, the step（1）In activation culture condition be：Shaking speed 100-160 r/min, temperature 20-
25 DEG C, light culture is activated 3-5 days.
Further, step（2）Described in separation be that 5-10 min are centrifuged under the conditions of 15000r/min.
Further, the step（3）The middle method for extracting Cordyceps militaris exocellular polysaccharide, step is as follows：
Take step（2）Obtained zymotic fluid, adds 95 % ethanol of 4 times of volumes, and 4 DEG C stand 5 h, 12000 rpm after mixing
10 min are centrifuged, precipitation is taken, is 1 M NaOH solution, 60 DEG C of 1 h of dissolving with 6 mL concentration, Cordyceps militaris exocellular polysaccharide is made.
Further, the step（3）The middle method for extracting Cordyceps militaris intracellular polyse, step is as follows：
By step（2）Obtained 65 DEG C of drying of cordyceps mycelium, smash, add 6mL by every gram of cordyceps mycelium powder
Concentration is 1 M NaOH solution, and 60 DEG C extract 2 h, and then 12000 r/min centrifuge 5 min, take supernatant, and pupa is made
Cordyceps sinensis intracellular polyse.
Marine microalgae as the original and highly important living marine resources of a class, rich in polyunsaturated fatty acid, polysaccharide,
The various bioactivators such as polypeptide, now mainly for the preparation of bioenergy and aquaculture feed, on the work for fermentation
Property material prepare report it is more rare.It is the microalgae that a class is rich in polyunsaturated fatty acid to split kettle algae, is often used to production high
Purity docosahexaenoic acid（Docosahexaenoic acid, DHA）Health products, infant's dairy products additive etc..Split
Algae-residue its protein content produced by after the extracted polyunsaturated fatty acid of kettle algae may be up to more than 40%, however at present these
Algae-residue is treated as feed or soil fertility quality mostly, and protein resource is not fully developed and utilized.The production of kettle algae fermentation protein will be split
The liquid fermentation that thing is applied to Cordyceps militaris is produced, domestic to improve the secondary metabolites such as Cordyceps sinensis polysaccharide yield production quantifier elimination
It is outer that there is not been reported.
The beneficial effects of the present invention are：
1st, the present invention will be split liquid fermentation of the kettle algae fermentation protein product applied to Polysaccharides in Cultured Cordyceps militaris and be produced, and intracellular polyse yield can
Up to 2.31g/L, yield of extracellular polysaccharide improves 1.9 times and more than 7.6 times than prior art respectively up to 8.37g/L；This hair
The output of the bright Cordyceps sinensis polysaccharide active ingredient for substantially increasing overall Cordyceps militaris, with good industrial applications prospect；
2nd, the present invention uses liquid aerobic fermentation method, and generally 5-7 days, relative to prior art, yield was high, and the cycle is short, nothing
The process such as quiescent culture and induction synthetic product is needed, production efficiency is higher, and the Polysaccharides in Cultured Cordyceps militaris produced can be directly used for being immunized
The preparation of the medicine such as power regulation, antitumor, hypoglycemic；
3rd, extensively, cost is relatively low, and preparation method is also relatively easy for the raw material sources of the present invention for splitting kettle algae fermentation protein product,
Can scale extract production, have to the fermenting and producing of Cordyceps militaris Cordyceps sinensis polysaccharide class material and promote well and lifting effect；
4th, Cordyceps militaris zymotechnique of the present invention is simple, and environment-protecting and non-poisonous, the cost of raw material is cheap, and full fermentation
Process control, is not limited by external environment condition, is adapted to industrialized production and popularization and application.
5th, the formula of liquid fermentation medium and seed fermentation culture medium is optimized the present invention, can be significantly
Improve the yield of Polysaccharides in Cultured Cordyceps militaris.
Embodiment
Technical scheme is clearly and completely described below in conjunction with the embodiment of the present invention, it is clear that retouched
The embodiment stated is only a part of embodiment of the invention, rather than whole embodiments.Based on the embodiment in the present invention, sheet
The every other embodiment that field those of ordinary skill is obtained under the premise of creative work is not made, belongs to the present invention
The scope of protection.
Embodiment 1
A kind of fermentation process of high concentration Cordyceps sinensis polysaccharide, comprises the following steps：
（1）Cordyceps militaris spawn is inoculated into PD liquid fermentation mediums, activation culture is carried out, then by percentage by volume 15%
Inoculum concentration be connected in seed fermentation culture medium, carry out seed culture, seed culture condition is：25 DEG C of temperature, shaking table culture 2
My god, obtain seed liquor；
（2）By step（1）Seed liquor is made to be inoculated in liquid fermentation medium by 10 % volume ratios, in 25 DEG C of bar of temperature
Under part, then liquid fermentation and culture 1 day adds active-fermented broth, and the concentration for making to split kettle algae fermentation protein product is 0.85 mg/
ML, then proceedes to culture 2 days, and separation obtains cordyceps mycelium and zymotic fluid；
（3）From step（2）Cordyceps militaris exocellular polysaccharide is extracted in obtained zymotic fluid, from step（2）Obtained cordyceps mycelium
Middle extraction Cordyceps militaris intracellular polyse, mixing Cordyceps militaris exocellular polysaccharide and Cordyceps militaris intracellular polyse, produce Cordyceps sinensis polysaccharide；
The step（1）In seed fermentation culture medium, every liter of component is as follows：
3 g glucose, 1.5 g yeast powder supernatants, 0.1 g MgSO4,0.03 g CaCl2,0.1 g KH2PO4,0.1
GK2HPO4,2-5 diameter 3-6 mm bead；
The step（2）In liquid fermentation medium, every liter of component is as follows：
3 g lactose, 2 g molasses, 2 g bean cake powders, 1.5 g peptones, 0.1 g MgSO4,0.03 g CaCl2,0.1 g
KH2PO4、0.1 g K2HPO4 ；
The active-fermented broth is splits kettle algae tunning, and its preparation method is：
Over cleaning of learning from else's experience splits kettle algae raw material, sets enzymatic hydrolysis condition as solid-liquid ratio 1: 10（ m/V）, cellulase Cellulase
ACCF-4740 additions 2%, 55 DEG C of temperature, pH 4.5, the h of reaction time 1.3, are digested, cellulose hydrolyzation terminates
Afterwards, boiling water bath goes out enzyme 15 min, and 9000 r/min centrifugations collect supernatant and obtain splitting kettle algae cellulose hydrolyzation liquid；Split kettle algae
Enzymolysis liquid goes out after enzyme activity, regulation pH value to 6.2-6.6, high pressure steam sterilization（121 DEG C, 20 min）, then it is inoculated with volume fraction
The Pediococcus pentosaceus liquid of 1% MRS culture mediums activation stands, fermented to kettle algae enzymolysis liquid, 37 DEG C of constant incubators are split；
After fermentation ends, 9000 r/min centrifugations collect supernatant and obtain splitting kettle algae tunning, the active-fermented broth as prepared.
Further, the step（1）In activation culture condition be：The r/min of shaking speed 160,25 DEG C of temperature, secretly
Culture activation 3 days.
Further, step（2）Described in separation be that 5 min are centrifuged under the conditions of 15000r/min.
Further, the step（3）The middle method for extracting Cordyceps militaris exocellular polysaccharide, step is as follows：
Take step（2）Obtained zymotic fluid, adds 95 % ethanol of 4 times of volumes, and 4 DEG C stand 5 h, 12000 rpm after mixing
10 min are centrifuged, precipitation is taken, is 1 M NaOH solution, 60 DEG C of 1 h of dissolving with 6 mL concentration, Cordyceps militaris exocellular polysaccharide is made.
Further, the step（3）The middle method for extracting Cordyceps militaris intracellular polyse, step is as follows：
By step（2）Obtained 65 DEG C of drying of cordyceps mycelium, smash, add 6mL by every gram of cordyceps mycelium powder
Concentration is 1 M NaOH solution, and 60 DEG C extract 2 h, and then 12000 r/min centrifuge 5 min, take supernatant, and pupa is made
Cordyceps sinensis intracellular polyse.
A kind of fermentation process for high concentration Cordyceps sinensis polysaccharide that the present embodiment is provided, greatly optimizes Polysaccharides in Cultured Cordyceps militaris fermentation
Process conditions, effectively shorten fermentation period and production cost, improve the worm that overall production efficiency and fermenting and producing go out
The concentration of grass polysaccharide, is that the offer of the fermenting and producing of modern industrialization Cordyceps sinensis polysaccharide is provided powerful support for.
Embodiment 2
A kind of fermentation process of high concentration Cordyceps sinensis polysaccharide, comprises the following steps：
（1）Cordyceps militaris spawn is inoculated into PD liquid fermentation mediums, activation culture is carried out, then by percentage by volume 10%
Inoculum concentration be connected in seed fermentation culture medium, carry out seed culture, seed culture condition is：20 DEG C of temperature, shaking table culture 2
My god, obtain seed liquor；
（2）By step（1）Seed liquor is made to be inoculated in liquid fermentation medium by 5 % volume ratios, in 20 DEG C of condition of temperature
Under, then liquid fermentation and culture 6 days adds active-fermented broth, and the concentration for making to split kettle algae fermentation protein product is 0.85 mg/
ML, then proceedes to culture 2-9 days, and separation obtains cordyceps mycelium and zymotic fluid；
（3）From step（2）Cordyceps militaris exocellular polysaccharide is extracted in obtained zymotic fluid, from step（2）Obtained cordyceps mycelium
Middle extraction Cordyceps militaris intracellular polyse, mixing Cordyceps militaris exocellular polysaccharide and Cordyceps militaris intracellular polyse, produce Cordyceps sinensis polysaccharide；
The step（1）In seed fermentation culture medium, every liter of component is as follows：
3 g glucose, 1.5 g yeast powder supernatants, 0.1 g MgSO4,0.03 g CaCl2,0.1 g KH2PO4,0.1
GK2HPO4,2-5 diameter 3-6 mm bead；
The step（2）In liquid fermentation medium, every liter of component is as follows：
3 g lactose, 2 g molasses, 2 g bean cake powders, 1.5 g peptones, 0.1 g MgSO4,0.03 g CaCl2,0.1 g
KH2PO4、0.1 g K2HPO4 ；
The active-fermented broth is splits kettle algae tunning, and its preparation method is：
Over cleaning of learning from else's experience splits kettle algae raw material, sets enzymatic hydrolysis condition as solid-liquid ratio 1: 10（ m/V）, cellulase Cellulase
ACCF-4740 additions 2%, 55 DEG C of temperature, pH 4.5, the h of reaction time 1.3, are digested, cellulose hydrolyzation terminates
Afterwards, boiling water bath goes out enzyme 15 min, and 9000 r/min centrifugations collect supernatant and obtain splitting kettle algae cellulose hydrolyzation liquid；Split kettle algae
Enzymolysis liquid goes out after enzyme activity, regulation pH value to 6.2-6.6, high pressure steam sterilization（121 DEG C, 20 min）, then it is inoculated with volume fraction
The Pediococcus pentosaceus liquid of 1% MRS culture mediums activation stands, fermented to kettle algae enzymolysis liquid, 37 DEG C of constant incubators are split；
After fermentation ends, 9000 r/min centrifugations collect supernatant and obtain splitting kettle algae tunning, the active-fermented broth as prepared.
Further, the step（1）In activation culture condition be：Shaking speed 100r/min, 25 DEG C of temperature, secretly
Culture activation 5 days.
Further, step（2）Described in separation be that 10 min are centrifuged under the conditions of 15000r/min.
Further, the step（3）The middle method for extracting Cordyceps militaris exocellular polysaccharide, step is as follows：
Take step（2）Obtained zymotic fluid, adds 95 % ethanol of 4 times of volumes, and 4 DEG C stand 5 h, 12000 rpm after mixing
10 min are centrifuged, precipitation is taken, is 1 M NaOH solution, 60 DEG C of 1 h of dissolving with 6 mL concentration, Cordyceps militaris exocellular polysaccharide is made.
Further, the step（3）The middle method for extracting Cordyceps militaris intracellular polyse, step is as follows：
By step（2）Obtained 65 DEG C of drying of cordyceps mycelium, smash, add 6mL by every gram of cordyceps mycelium powder
Concentration is 1 M NaOH solution, and 60 DEG C extract 2 h, and then 12000 r/min centrifuge 5 min, take supernatant, and pupa is made
Cordyceps sinensis intracellular polyse.
Embodiment 3
A kind of fermentation process of high concentration Cordyceps sinensis polysaccharide, comprises the following steps：
（1）Cordyceps militaris spawn is inoculated into PD liquid fermentation mediums, activation culture is carried out, then by percentage by volume
12.7% inoculum concentration is connected in seed fermentation culture medium, carries out seed culture, and seed culture condition is：23 DEG C of temperature, shaking table
Culture 3 days, obtains seed liquor；
（2）By step（1）Seed liquor is made to be inoculated in liquid fermentation medium by 7 % volume ratios, in 22 DEG C of condition of temperature
Under, then liquid fermentation and culture 3 days adds active-fermented broth, and the concentration for making to split kettle algae fermentation protein product is 0.5 mg/mL,
Culture 4 days is then proceeded to, separation obtains cordyceps mycelium and zymotic fluid；
（3）From step（2）Cordyceps militaris exocellular polysaccharide is extracted in obtained zymotic fluid, from step（2）Obtained cordyceps mycelium
Middle extraction Cordyceps militaris intracellular polyse, mixing Cordyceps militaris exocellular polysaccharide and Cordyceps militaris intracellular polyse, produce Cordyceps sinensis polysaccharide；
The step（1）In seed fermentation culture medium, every liter of component is as follows：
3 g glucose, 1.5 g yeast powder supernatants, 0.1 g MgSO4,0.03 g CaCl2,0.1 g KH2PO4,0.1
GK2HPO4,2-5 diameter 3-6 mm bead；
The step（2）In liquid fermentation medium, every liter of component is as follows：
3 g lactose, 2 g molasses, 2 g bean cake powders, 1.5 g peptones, 0.1 g MgSO4,0.03 g CaCl2,0.1 g
KH2PO4、0.1 g K2HPO4 ；
The active-fermented broth is splits kettle algae tunning, and its preparation method is：
Over cleaning of learning from else's experience splits kettle algae raw material, sets enzymatic hydrolysis condition as solid-liquid ratio 1: 10（ m/V）, cellulase Cellulase
ACCF-4740 additions 2%, 55 DEG C of temperature, pH 4.5, the h of reaction time 1.3, are digested, cellulose hydrolyzation terminates
Afterwards, boiling water bath goes out enzyme 15 min, and 9000 r/min centrifugations collect supernatant and obtain splitting kettle algae cellulose hydrolyzation liquid；Split kettle algae
Enzymolysis liquid goes out after enzyme activity, regulation pH value to 6.2-6.6, high pressure steam sterilization（121 DEG C, 20 min）, then it is inoculated with volume fraction
The Pediococcus pentosaceus liquid of 1% MRS culture mediums activation stands, fermented to kettle algae enzymolysis liquid, 37 DEG C of constant incubators are split；
After fermentation ends, 9000 r/min centrifugations collect supernatant and obtain splitting kettle algae tunning, the active-fermented broth as prepared.
Further, the step（1）In activation culture condition be：Shaking speed 125r/min, 22 DEG C of temperature, dark training
Supporting 4 days.
Further, step（2）Described in separation be that 7 min are centrifuged under the conditions of 15000r/min.
Further, the step（3）The middle method for extracting Cordyceps militaris exocellular polysaccharide, step is as follows：
Take step（2）Obtained zymotic fluid, adds 95 % ethanol of 4 times of volumes, and 4 DEG C stand 5 h, 12000 rpm after mixing
10 min are centrifuged, precipitation is taken, is 1 M NaOH solution, 60 DEG C of 1 h of dissolving with 6 mL concentration, Cordyceps militaris exocellular polysaccharide is made.
Further, the step（3）The middle method for extracting Cordyceps militaris intracellular polyse, step is as follows：
By step（2）Obtained 65 DEG C of drying of cordyceps mycelium, smash, add 6mL by every gram of cordyceps mycelium powder
Concentration is 1 M NaOH solution, and 60 DEG C extract 2 h, and then 12000 r/min centrifuge 5 min, take supernatant, and pupa is made
Cordyceps sinensis intracellular polyse.
It is obvious to a person skilled in the art that the invention is not restricted to the details of above-mentioned one exemplary embodiment, Er Qie
In the case of without departing substantially from spirit or essential attributes of the invention, the present invention can be realized in other specific forms.Therefore, no matter
From the point of view of which point, embodiment all should be regarded as exemplary, and be nonrestrictive, the scope of the present invention is by appended power
Profit is required rather than described above is limited, it is intended that all in the implication and scope of the equivalency of claim by falling
Change is included in the present invention.
Moreover, it will be appreciated that although the present specification is described in terms of embodiments, not each embodiment is only wrapped
Containing an independent technical scheme, this narrating mode of specification is only that for clarity, those skilled in the art should
Using specification as an entirety, the technical solutions in the various embodiments may also be suitably combined, forms those skilled in the art
It may be appreciated other embodiment.The ins and outs not being described in detail in the present invention, can pass through any in this area
Prior art is realized.Particularly, all technical characterstics not being described in detail can be realized by any prior art in the present invention.
Claims (5)
1. a kind of fermentation process of high concentration Cordyceps sinensis polysaccharide, it is characterised in that comprise the following steps：
（1）Cordyceps militaris spawn is inoculated into PD liquid fermentation mediums, activation culture is carried out, then by percentage by volume 10-
15% inoculum concentration is connected in seed fermentation culture medium, carries out seed culture, and seed culture condition is：20-25 DEG C of temperature, shaking table
Culture 2-4 days, obtains seed liquor；
（2）By step（1）Seed liquor is made to be inoculated in liquid fermentation medium by 5-10 % volume ratios, in 20-25 DEG C of temperature
Under conditions of, then liquid fermentation and culture 1-6 days adds active-fermented broth, makes the concentration for splitting kettle algae fermentation protein product be
0.15-0.85 mg/mL, then proceed to culture 2-9 days, and separation obtains cordyceps mycelium and zymotic fluid；
（3）From step（2）Cordyceps militaris exocellular polysaccharide is extracted in obtained zymotic fluid, from step（2）Obtained cordyceps mycelium
Middle extraction Cordyceps militaris intracellular polyse, mixing Cordyceps militaris exocellular polysaccharide and Cordyceps militaris intracellular polyse, produce Cordyceps sinensis polysaccharide；
The step（1）In seed fermentation culture medium, every liter of component is as follows：
3 g glucose, 1.5 g yeast powder supernatants, 0.1 g MgSO4,0.03 g CaCl2,0.1 g KH2PO4,0.1
GK2HPO4,2-5 diameter 3-6 mm bead；
The step（2）In liquid fermentation medium, every liter of component is as follows：
3 g lactose, 2 g molasses, 2 g bean cake powders, 1.5 g peptones, 0.1 g MgSO4,0.03 g CaCl2,0.1 g
KH2PO4、0.1 g K2HPO4 ；
The active-fermented broth is splits kettle algae tunning, and its preparation method is：
Over cleaning of learning from else's experience splits kettle algae raw material, sets enzymatic hydrolysis condition as solid-liquid ratio 1: 10（ m/V）, cellulase Cellulase
ACCF-4740 additions 2%, 55 DEG C of temperature, pH 4.5, the h of reaction time 1.3, are digested, cellulose hydrolyzation terminates
Afterwards, boiling water bath goes out enzyme 15 min, and 9000 r/min centrifugations collect supernatant and obtain splitting kettle algae cellulose hydrolyzation liquid；Split kettle algae
Enzymolysis liquid goes out after enzyme activity, regulation pH value to 6.2-6.6, high pressure steam sterilization（121 DEG C, 20 min）, then it is inoculated with volume fraction
The Pediococcus pentosaceus liquid of 1% MRS culture mediums activation stands, fermented to kettle algae enzymolysis liquid, 37 DEG C of constant incubators are split；
After fermentation ends, 9000 r/min centrifugations collect supernatant and obtain splitting kettle algae tunning, the active-fermented broth as prepared.
2. the fermentation process of the high concentration Cordyceps sinensis polysaccharide as described in claim 1, it is characterised in that the step（1）In
Activation culture condition is：Shaking speed 100-160 r/min, 20-25 DEG C of temperature, light culture is activated 3-5 days.
3. the fermentation process of the high concentration Cordyceps sinensis polysaccharide as described in claim 1, it is characterised in that step（2）Described in
Separation is that 5-10 min are centrifuged under the conditions of 15000r/min.
4. the fermentation process of the high concentration Cordyceps sinensis polysaccharide as described in claim 1, it is characterised in that the step（3）In carry
The method for taking Cordyceps militaris exocellular polysaccharide, step is as follows：
Take step（2）Obtained zymotic fluid, adds 95 % ethanol of 4 times of volumes, and 4 DEG C stand 5 h, 12000 rpm after mixing
10 min are centrifuged, precipitation is taken, is 1 M NaOH solution, 60 DEG C of 1 h of dissolving with 6 mL concentration, Cordyceps militaris exocellular polysaccharide is made.
5. the fermentation process of the high concentration Cordyceps sinensis polysaccharide as described in claim 1, it is characterised in that the step（3）In carry
The method for taking Cordyceps militaris intracellular polyse, step is as follows：
By step（2）Obtained 65 DEG C of drying of cordyceps mycelium, smash, add 6mL by every gram of cordyceps mycelium powder
Concentration is 1 M NaOH solution, and 60 DEG C extract 2 h, and then 12000 r/min centrifuge 5 min, take supernatant, and pupa is made
Cordyceps sinensis intracellular polyse.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710355633.8A CN106967765A (en) | 2017-05-19 | 2017-05-19 | The fermentation process of high concentration Cordyceps sinensis polysaccharide |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710355633.8A CN106967765A (en) | 2017-05-19 | 2017-05-19 | The fermentation process of high concentration Cordyceps sinensis polysaccharide |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN106967765A true CN106967765A (en) | 2017-07-21 |
Family
ID=59325902
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201710355633.8A Pending CN106967765A (en) | 2017-05-19 | 2017-05-19 | The fermentation process of high concentration Cordyceps sinensis polysaccharide |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN106967765A (en) |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS5668389A (en) * | 1979-11-05 | 1981-06-09 | Yoshio Tani | Culture of mycelium type yeast and fermentative production of high-molecular substance |
| CN101353382A (en) * | 2008-09-11 | 2009-01-28 | 浙江工业大学 | Extraction method of antioxidative active Cordyceps sinensis polysaccharide |
| CN102618598A (en) * | 2012-04-16 | 2012-08-01 | 山东省农业科学院农业资源与环境研究所 | Liquid fermentation method for improving yield of cordyceps sinensis polysaccharide by utilizing expansin |
| CN102715296A (en) * | 2012-06-13 | 2012-10-10 | 镇江市食用菌研究所 | Worm grass tea and making method thereof |
-
2017
- 2017-05-19 CN CN201710355633.8A patent/CN106967765A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS5668389A (en) * | 1979-11-05 | 1981-06-09 | Yoshio Tani | Culture of mycelium type yeast and fermentative production of high-molecular substance |
| CN101353382A (en) * | 2008-09-11 | 2009-01-28 | 浙江工业大学 | Extraction method of antioxidative active Cordyceps sinensis polysaccharide |
| CN102618598A (en) * | 2012-04-16 | 2012-08-01 | 山东省农业科学院农业资源与环境研究所 | Liquid fermentation method for improving yield of cordyceps sinensis polysaccharide by utilizing expansin |
| CN102715296A (en) * | 2012-06-13 | 2012-10-10 | 镇江市食用菌研究所 | Worm grass tea and making method thereof |
Non-Patent Citations (2)
| Title |
|---|
| 张丽姣: "海带酶解液的戊糖片球菌发酵及其产物的抗氧化性、抑菌性", 《食品科学》 * |
| 汪少芸等: "海洋微藻裂壶藻发酵液胞外多糖的制备及活性研究", 《福州大学学报（自然科学版）》 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN101543285B (en) | Wheat-bran dietary fiber composite functional fungus powder product and preparation process thereof | |
| CN103190576B (en) | Cordyceps militaris soybean and preparation method thereof | |
| CN104323222B (en) | A kind of bacterium mushroom powder being rich in farnoquinone and preparation method thereof | |
| CN103483040B (en) | Chinese caterpillar fungus pilot scale liquid submerged fermentation substratum and fermentation method for producing thereof | |
| CN104893992A (en) | Submerged fermentation method for ganoderma lucidum | |
| CN104846018A (en) | Cordyceps enzyme deep layer liquid fermentation and expanding culture method | |
| CN110507681A (en) | A kind of technique of higher value application flower of Panax ginseng | |
| CN102599420B (en) | Edible mushroom and fungus composite-flavor nutrient noodles prepared through submerged fermentation of liquid and production method thereof | |
| CN104087632B (en) | A kind of deep layer liquid fermentation produces the method for Phellinus igniarius (L. ex Fr.) Quel. extracellular polysaccharide | |
| CN103421861A (en) | Method of manufacturing cordyceps sinensis (Berk.) sacc polysaccharide by liquid state fermentation of rice bran and bran complete feed | |
| CN103229666B (en) | Cordyceps militaris peanut and preparing method thereof | |
| CN101805760A (en) | Method for producing microbial oil comprising arachidonic acid | |
| CN105535035B (en) | A kind of Inonotus obliquus fermented and cultured composition and preparation method thereof | |
| CN103555786A (en) | Method for producing polysaccharides through liquid state fermentation of rice bran and wheat bran complete material by using ganoderma lucidum mutant strain | |
| CN106867907A (en) | A kind of overcompensation cultural method for improving heterotrophic microalgae protein content | |
| CN103160550A (en) | Preparation method of compound polysaccharide consisting of schizophyllum commune exopolysaccharide and oat polysaccharide | |
| CN109022501A (en) | A method of ferulic acid is obtained using waste | |
| CN106479899A (en) | A kind of cordyceps militaris link bacterial strain and its application in cordycepin is prepared | |
| CN102329833A (en) | Method for producing sophorose ester through fermenting waste molasses and waste glycerin | |
| CN106434377B (en) | A kind of cordyceps militaris link bacterial strain and its application in preparation production rouge Cordyceps militaris product | |
| CN102584363B (en) | Method for producing medical mycelium or medical and edible dual-purpose mycelium by using yellow wine lees as liquid medium | |
| CN106967765A (en) | The fermentation process of high concentration Cordyceps sinensis polysaccharide | |
| CN102199636A (en) | Efficient preparation process of gamma-amino-n-butyric acid | |
| CN102771854A (en) | Phellinus igniarius health-care beverage prepared through liquid submerged fermentation and preparation method thereof | |
| CN102041287B (en) | Method for producing polysaccharide by crossly culturing tremella fuciformis basidiospores by stages |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20170721 |
|
| RJ01 | Rejection of invention patent application after publication |